The biochemistry of IgE receptor-mediated activation of mast cells is n=being studied in two separate complementary projects which are summarized below. 1) Inhibition of IgE receptor-mediated exocytosis from mast cells by the immunosuppressive drugs cyclosporin A (CsA) and FK506. Immunosuppressive drugs such as CsA and FK506 are considered by many to specifically inhibit T cell functions; however, the same concentration of CsA that inhibits T cell receptor-mediated activation of T cells also inhibits IgE receptor-mediated degranulation of mast cells. These studies have been extended by studying the effect of FK506 and its structural analog, rapamycin, on IgE receptor-mediated degranulation of mast cells. The intracellular target for rapamycin and FK506 (FKBP) is distinct from the intracellular target for CsA (cyclophilin). Like CsA, FK506 inhibits degranulation of mast cells without effecting any other IgE receptor-mediated function; however, FK506 is 100 times more potent than CsA. The lC(50) for inhibition is 200 nM and 2 nM for CsA and FK506, respectively. Rapamycin and 506BD both compete with FK506 for binding to FKBP. Pretreatment of mast cells with either rapamycin or 506BD prevents the inhibition by FK506. Thus, it appears as though a FKBP-FK506 complex as well as a cyclophilin-CsA complex inhibits receptor-mediated activation of mast cells. 2) Activation of membrane-associated phospholipase C by aggregation of IgE receptors. Currently, little is known concerning the mechanism of activation of PLC and, in fact it is not even known which of the PLC isozymes present in mast cells is involved in receptor-mediated PI hydrolysis. A major obstacle to these studies is the lack of a cell-free system in which this activation can be studied. We have established a system in which aggregation of IgE receptors results in activation of membrane-associated PLC. Rat basophilic leukemia (RBL) cells are activated by aggregation of the IgE receptors. The PLC activity in membranes isolated from activated cells is increased by 150% above membranes from control cells. This system will be used to identify which PLC isozyme is activated and the mechanism of this activation.